Combinatorial anti-retroviral therapies have been remarkably effective in controlling the progression of AIDS and in prolonging the life of patients infected by HIV. However, while the current drug therapy can delay the progression of the disease it can not eradicate the virus, and its use readily leads to the emergence of drug resistant HIV strains. Furthermore, the emergence of viral resistance to one drug frequently results in a cross resistance to other drugs that exhibit similar chemical structures and mechanisms of action. Thus, the identification of novel viral targets and/or the development of new classes of antiviral compounds are essential in the fight against HIV/AIDS. HIV-1 reverse transcriptase (RT) is a heterodimeric enzyme consisting of a 66-kDa subunit (p66) and a p66- derived 51-kDa subunit (p51). The DNA polymerase and ribonuclease H (RNase H) activities of the enzyme are entirely dependent on the dimeric structure of the enzyme. Accordingly, dimerization of HIV-1 RT provides a novel therapeutic target for the identification of a new class of antiviral compounds. We have recently developed an in vitro high throughput screening (HTS) assay that reproducibly detects the intersubunit interactions between the p66 and p51 subunits of HIV-1 RT. Furthermore this assay is sensitive to compounds that have previously been shown to either inhibit or enhance the inter-subunit interactions of the enzyme. In light of this, the project described in this proposal comprises two Specific Aims: (1) to optimize and validate the in vitro HTS assay for HIV-1 RT dimerization; and (2) to screen a chemical and a natural product library for inhibitors of HIV-1 RT dimerization. Due to its essential role in HIV-1 replication, RT is already a primary target for the development of antiviral compounds. However, the elucidation and/or identification of inhibitors of HIV-1 RT dimerization would create a new therapeutic class of drugs that would exhibit mechanisms of action entirely different from the nucleoside and nonnucleoside RT inhibitors that are currently used in the treatment of HIV-1 infected individuals.